RESUMEN
BACKGROUND: The administration of peroxisome proliferator-activated receptor gamma (PPARgamma) agonists to low-density lipoprotein (LDL)-receptor-deficient mice resulted in a reduction in the atherosclerotic lesion area in male mice, but not in female mice. The male mice also exhibited reduction in insulin resistance while the female mice did not. To further examine the relationship between PPARgamma agonists, insulin resistance and atherosclerosis, we used the model of accelerated atherosclerosis in male apolipoprotein E (apoE)-deficient mice rendered diabetic by low-dose streptozotocin (STZ). METHODS: Male, apoE-deficient mice (n = 48) were randomly divided into four groups. To induce diabetes, two groups received low-dose STZ and two groups served as controls. After diabetes induction, rosiglitazone (a PPARgamma agonist) was administered by oral gavage to one of the diabetic and one of the non-diabetic groups. RESULTS: Rosiglitazone reduced significantly the atherosclerotic aortic plaque area in both diabetic and non-diabetic apoE-deficient mice: 340 +/- 54 vs. 201 +/- 27 micromol2 (p = 0.001) in diabetic mice; 243 +/- 22 vs. 158 +/- 27 micromol2 (p = 0.001) in non-diabetic mice. Also, rosiglitazone reduced the correlation coefficient between plasma glucose and the degree of atherosclerosis (p < 0.0025) without affecting plasma glucose levels. The rosiglitazone-treated mice, both diabetic and non-diabetic, had higher lipid levels. CONCLUSIONS: Rosiglitazone-treated animals showed less atherosclerosis despite higher lipid levels and similar glucose levels. These data suggest a direct anti-atherogenic effect of rosiglitazone on the arterial wall.
Asunto(s)
Enfermedades de la Aorta/tratamiento farmacológico , Arteriosclerosis/tratamiento farmacológico , Angiopatías Diabéticas/tratamiento farmacológico , Fibrinolíticos/uso terapéutico , Tiazoles/uso terapéutico , Tiazolidinedionas , Animales , Enfermedades de la Aorta/sangre , Enfermedades de la Aorta/patología , Apolipoproteínas E/deficiencia , Arteriosclerosis/sangre , Arteriosclerosis/patología , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Colesterol/sangre , Diabetes Mellitus Experimental/sangre , Angiopatías Diabéticas/sangre , Masculino , Ratones , Receptores Citoplasmáticos y Nucleares/agonistas , Rosiglitazona , Factores de Transcripción/agonistas , Triglicéridos/sangreRESUMEN
BACKGROUND: Human 15-lipoxygenase (LO) and its murine analogue 12/15-LO are capable of directly oxidizing esterified fatty acids in lipoproteins and phospholipids. Because these oxidized products possess atherogenic properties, it was suggested that LOs may be involved in enhancing atherogenesis. Previous in vivo tests of the role of LOs in atherogenesis animal models, however, have yielded conflicting results. METHODS AND RESULTS: Aiming to study the role of the 12/15-LO in murine atherogenesis, we crossed LDL-receptor-deficient mice (LDL-R(-/-)) with 12/15-LO-knockout mice and evaluated plaque formation 3 to 18 weeks after initiation of a high-fat diet. Atherosclerotic lesions were considerably reduced in the LDL-R/12/15-LO-double-knockout mice compared with LDL-R(-/-) mice at 3, 9, 12, and 18 weeks, at the aortic root as well as throughout the aorta. The cellular composition of plaques from mice deficient in 12/15-LO did not differ with respect to macrophage and T-lymphocyte content compared with plaques from 12/15-LO littermates. CONCLUSIONS: 12/15-LO plays a dominant role in promoting atherogenesis in LDL-R(-/-) mice.
Asunto(s)
Araquidonato 12-Lipooxigenasa/genética , Araquidonato 12-Lipooxigenasa/fisiología , Araquidonato 15-Lipooxigenasa/genética , Araquidonato 15-Lipooxigenasa/fisiología , Arteriosclerosis/etiología , Receptores de LDL/genética , Animales , Aorta/patología , Arteriosclerosis/sangre , Arteriosclerosis/patología , Colesterol/sangre , Dieta Aterogénica , Recuento de Leucocitos , Macrófagos , Ratones , Ratones Noqueados , Linfocitos T , Triglicéridos/sangreRESUMEN
15-Lipoxygenase (15-LOX) is a lipid-oxidizing enzyme that is involved in cell cycle regulation. To evaluate the effect of 15-LOX on reproduction, we studied transgenic mice that overexpress 15-LOX. The transgene was introduced over the genetic background of the low density lipoprotein receptor deficient mice (LDL-R(-/-)) and reproduction was compared to LDL-R(-/-) mice. We found a lower pregnancy rate in the 15-LOX/LDL-R(-/-) mice as compared to the LDL-R(-/-) mice (62.74 vs. 79.1%, p < 0.01). Additionally, a remarkably higher number of resorptions per pregnancy was found in the 15-LOX/LDL-R(-/-) mice (16.7 vs. 3.27%, p < 0.001) and it was accompanied by a significantly higher activity of the 15-LOX enzyme in these resorptions as compared to other tissues. These findings may implicate a role for 15-LOX in the development of spontaneous abortions.
Asunto(s)
Aborto Espontáneo/metabolismo , Araquidonato 15-Lipooxigenasa/metabolismo , Ratones Endogámicos C57BL/metabolismo , Ratones Transgénicos/metabolismo , Animales , Tasa de Natalidad , Femenino , Tamaño de la Camada , Ratones , Modelos Animales , Embarazo , Receptores de LDL/metabolismoRESUMEN
Gene therapy directed specifically to the vascular wall, particularly to angiogenic endothelial cells is a prerequisite in vascular disease treatment. Angiogenesis is a major feature in many pathological conditions including wound healing, solid tumors, developing metastases, ischemic heart diseases and diabetic retinopathy. In the present study we developed a tissue-specific gene therapy to the angiogenic blood vessels of tumor metastasis using an adeno-based vector containing the murine preproendothelin-1 (PPE-1) promoter. Genes activated by the PPE-1 promoter were highly expressed in bovine aortic endothelial cells in vitro. Systemic injection of the adenoviral vectors AdPPE-1-luciferase and AdCMV-luciferase to normal C57BL/6 mice, resulted in higher activity of PPE-1 promoter compared with CMV promoter in the aorta and vascularized tissues such as heart, kidney, lung and pancreas. Systemic administration of the adenoviral vector, in mice bearing Lewis lung carcinoma, resulted in high and specific activity of PPE-1 in the new vasculature of primary tumors and lung metastasis. Cellular distribution of the delivered gene revealed highest expression of GFP in angiogenic endothelial cells of the metastasis. We expect that this approach of 'vascular-directed' gene therapy will be applicable to both vascular diseases and cancer.
Asunto(s)
Carcinoma Pulmonar de Lewis/secundario , Endotelinas/genética , Endotelio Vascular/metabolismo , Terapia Genética/métodos , Neoplasias Pulmonares/secundario , Neovascularización Patológica , Precursores de Proteínas/genética , Adenoviridae/genética , Análisis de Varianza , Animales , Aorta , Carcinoma Pulmonar de Lewis/irrigación sanguínea , Carcinoma Pulmonar de Lewis/terapia , Bovinos , Células Cultivadas , Endotelina-1/genética , Expresión Génica , Marcación de Gen/métodos , Vectores Genéticos/administración & dosificación , Proteínas Fluorescentes Verdes , Hígado/metabolismo , Proteínas Luminiscentes/genética , Neoplasias Pulmonares/irrigación sanguínea , Neoplasias Pulmonares/terapia , Masculino , Ratones , Ratones Endogámicos C57BL , Microscopía Fluorescente , Microscopía de Contraste de Fase , Regiones Promotoras Genéticas , Estadísticas no ParamétricasRESUMEN
OBJECTIVE: The aim of the present study was to compare the accumulation of (125)I-labeled low-density lipoproteins (LDL), oxidized LDL (oxLDL), HDL and BSA in advanced atherosclerotic lesions of apoE-deficient mice. METHODS: (125)I-lipoproteins or (125)I-BSA were injected into the tail vein of apoE-deficient mice. Blood clearance of (125)I-lipoproteins and (125)I-BSA and their accumulation in atherosclerotic lesions were assayed. RESULTS: Blood clearance of (125)I-LDL and (125)I-HDL was moderate, and approximately 30% of the injected lipoproteins were present in plasma 24 h following injection. oxLDL was removed much faster from plasma, and less than 10% of (125)I-oxLDL was present in the circulation 30 min after (125)I-oxLDL injection. The clearance of (125)I-BSA from the circulation was slower than the lipoprotein clearance. The highest accumulation of LDL, oxLDL, HDL and BSA was detected in atherosclerotic lesions in the aortic arch and abdominal aorta, while lower accumulation was detected in the less atherosclerotic descending thoracic aorta. CONCLUSION: Our findings demonstrate that both (125)I-HDL and (125)I-BSA as well as (125)I-LDL are accumulated in atherosclerotic plaques and that they can be used for the detection of atherosclerotic lesions.
Asunto(s)
Aorta/diagnóstico por imagen , Arteriosclerosis/diagnóstico por imagen , Radioisótopos de Yodo , Lipoproteínas , Albúmina Sérica Bovina , Animales , Aorta/patología , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Arteriosclerosis/genética , Arteriosclerosis/patología , Modelos Animales de Enfermedad , Humanos , Radioisótopos de Yodo/farmacocinética , Lipoproteínas/farmacocinética , Lipoproteínas HDL/farmacocinética , Lipoproteínas LDL/farmacocinética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Cintigrafía , Albúmina Sérica Bovina/farmacocinéticaRESUMEN
To study the possible role of the human lipid-oxidizing enzyme 15-lipoxygenase (15-LO) in atherosclerosis, we overexpressed it specifically in the vascular wall of C57B6/SJL mice by using the murine preproendothelin-1 promoter. The mice overexpressing 15-LO were crossbred with low density lipoprotein (LDL) receptor-deficient mice to investigate atherogenesis. High levels of 15-LO were expressed in the atherosclerotic lesion in the double-transgenic mice as assessed by immunohistochemistry. The double-transgenic, 15-LO-overexpressing, LDL receptor-deficient mice (LDLR-/-/15LO) developed significantly larger atherosclerotic lesions at the aortic sinus compared with lesions in the LDL receptor-deficient (LDLR-/-) mice after 3 and 6 weeks (107,000 versus 28,000 microm(2) [P:<0.001] and 121,000 versus 87,000 microm(2) [P:<0.05], respectively) of an atherogenic diet. LDL from the LDLR-/-/15LO mice was more susceptible to oxidation than was the LDL from the control LDLR-/- mice, as shown by a shorter lag period for copper-induced conjugated diene formation. On the other hand, no differences were found in the levels of serum anti-oxidized LDL antibodies between the study groups. There were also no differences with respect to the density of macrophages and T lymphocytes infiltrating the lesions in both experimental groups. Taken together, these results support the hypothesis that 15-LO overexpression in the vessel wall is associated with enhanced atherogenesis.
Asunto(s)
Araquidonato 15-Lipooxigenasa/fisiología , Arteriosclerosis/enzimología , Endotelio Vascular/enzimología , Receptores de LDL/metabolismo , Animales , Anticuerpos/sangre , Araquidonato 15-Lipooxigenasa/biosíntesis , Endotelina-1 , Endotelinas/genética , Lípidos/sangre , Lipoproteínas/metabolismo , Lipoproteínas LDL/inmunología , Ratones , Ratones Transgénicos , Oxidación-Reducción , Precursores de Proteínas/genética , Receptores de LDL/deficienciaRESUMEN
OBJECTIVE: Magnesium (Mg) has previously been found to modulate blood lipid levels, atherogenesis and atherosclerosis in rabbits when used as a dietary supplement. In addition, we have reported that Mg fortification of drinking water can attenuate atherogenesis in male low-density lipoprotein (LDL)-receptor-deficient mice, but had a mild and nonsignificant effect on female mice fed a high-cholesterol diet supplemented with cholic acid. The aim of this study was to examine whether Mg has an antiatherogenic effect in female mice fed a high-cholesterol diet without cholic acid. METHODS: Two groups of female LDL-receptor-deficient mice were included. The mice received either distilled water or water with 50 g of Mg sulfate per liter. In the first (12 weeks) and second (6 weeks) stages of the experiment, the mice received low- and high-cholesterol diets, respectively, both without cholic acid. At the end of each stage of the experiment, blood was drawn for the determination of plasma Mg, calcium and lipid levels. In addition, the extent of atherosclerosis was determined at the aortic sinus level. RESULTS: Mg fortification was associated with higher levels of plasma Mg while the mice were on a high-cholesterol diet, and the extent of atherosclerosis at the aortic sinus was significantly decreased in the female mice that received high levels of Mg compared with the female mice that received distilled water. The female mice that received water fortified with Mg had lower levels of triglycerides after stage 2, whereas no differences regarding cholesterol levels were found. CONCLUSION: These results confirm that Mg fortification of drinking water is capable of inhibiting atherogenesis also in female LDL-receptor-deficient mice fed a high-cholesterol diet, and demonstrate the importance of the nutritional composition of diet in this experimental model.
Asunto(s)
Arteriosclerosis/prevención & control , Alimentos Fortificados , Magnesio/administración & dosificación , Receptores de LDL/deficiencia , Animales , Arteriosclerosis/etiología , Arteriosclerosis/patología , Calcio/sangre , Colesterol en la Dieta/efectos adversos , Dieta , Femenino , Lípidos/sangre , Magnesio/sangre , Ratones , Ratones Noqueados , Seno Aórtico/patología , Abastecimiento de AguaRESUMEN
Diabetes and atherosclerosis have been proposed to be influenced by immune and autoimmune mechanisms. A common incriminated antigen in both disorders is the heat shock protein (HSP)-60/65. In the current study, we established a model combining hyperglycemia with hyperlipidemia in LDL receptor-deficient (LDL-RD) mice and assessed its possible influences on lipid profile, HSP60/65, and atherogenesis. LDL-RD mice were injected either with streptozotocin to induce hyperglycemia or with citrate buffer (control). When hyperglycemia was induced, both study groups were challenged with a high-fat (Western) diet for 6 weeks. Plasma fasting glucose, lipid profile, and antibody levels to HSP65 and oxidized LDL were assessed. At death, the spleens from both groups were evaluated for their proliferative response to HSP65 and the consequent cytokine production. The extent of atherosclerosis was assessed at the aortic sinus. Plasma glucose, cholesterol, and triglyceride levels were elevated in mice injected with streptozotocin compared with control mice. Atherosclerotic lesions were significantly larger in the streptozotocin-injected hyperglycemic LDL-RD mice (132 +/- 23 x 10(5) microm2) in comparison to their normoglycemic litter-mates (20 +/- 6.6 x 10(5) microm2; P < 0.0001). Both humoral and cellular immune response to HSP65 was more pronounced in streptozotocin-injected mice. When challenged with HSP65 in vitro, splenocytes from streptozotocin-injected mice favored the production of the T-helper (TH)-1 cytokine gamma-interferon. In conclusion, we have established a mouse model that combines hyperglycemia with diet-induced hyperlipidemia in LDL-RD mice and studied its effect on atherosclerosis progression. The accelerated atherosclerotic process is associated with heightened immune response to HSP65 and a shift to a TH1 cytokine profile.
Asunto(s)
Arteriosclerosis/complicaciones , Proteínas Bacterianas , Hiperglucemia/complicaciones , Hiperlipidemias/complicaciones , Receptores de LDL/deficiencia , Animales , Anticuerpos/análisis , Aorta/patología , Arteriosclerosis/patología , Glucemia/análisis , Peso Corporal , Chaperonina 60 , Chaperoninas/inmunología , Citocinas/biosíntesis , Femenino , Hiperglucemia/sangre , Hiperglucemia/inducido químicamente , Hiperglucemia/inmunología , Hiperlipidemias/sangre , Hiperlipidemias/inducido químicamente , Hiperlipidemias/inmunología , Inmunidad , Inmunidad Celular , Lípidos/sangre , Lipoproteínas LDL/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Bazo/inmunología , Bazo/metabolismo , Bazo/patología , EstreptozocinaRESUMEN
OBJECTIVE: The renin-angiotensin system may play a role in the development of atherosclerosis. Nevertheless, different results from studies attempting to attenuate the process by inhibiting the converting enzyme were equivocal, and in those who succeeded, blood pressure was lowered and/or the lipid profile was improved in addition to the inhibition of the renin-angiotensin axis. The aim of this study is to investigate the effect of low doses of fosinopril, a converting enzyme inhibitor, on the development of atherosclerosis in LDL-receptor-deficient mice. METHODS: Three groups of 15 mice were fed a high-fat, high-cholesterol western diet. The three study groups received either distilled water (control group), or water supplemented with fosinopril 0.01 mg/kg/day (low-dose group) or with 0.1 mg/kg/day (high-dose group). Plasma aldosterone levels and lipid profiles were measured at the beginning and at the end of the study. After 10 weeks, the mice were sacrificed and the extent of atherosclerosis was assessed at the aortic sinus. RESULTS: Plasma aldosterone levels did not change in the control group, but decreased significantly in both treated groups from 74.7 to 39.3 ng/ml in the low-dose group (p < 0.003) and from 70.7 to 33.6 ng/ml in the high-dose group (p < 0.001). The lipid profile at the end of the study showed significantly lower levels of cholesterol and triglycerides in the high-dose group as compared to the low-dose group (p < 0.05). There was no difference between the three groups regarding the area of atherosclerosis at the aortic sinus: 157,000 +/- 34,000, 130,000 +/- 58,000 and 145,000 +/- 26,000 microm(2) in the control, low-dose and high-dose groups, respectively. CONCLUSION: Inhibition of the renin-angiotensin-aldosterone axis by itself does not prevent the development of early atherosclerosis in LDL-receptor-deficient mice.
Asunto(s)
Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Arteriosclerosis/prevención & control , Fosinopril/uso terapéutico , Receptores de LDL/deficiencia , Sistema Renina-Angiotensina/fisiología , Aldosterona/sangre , Animales , Arteriosclerosis/sangre , Arteriosclerosis/inducido químicamente , Arteriosclerosis/patología , Peso Corporal/efectos de los fármacos , Colesterol/sangre , Colesterol en la Dieta/efectos adversos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de LDL/genética , Seno Aórtico/efectos de los fármacos , Seno Aórtico/patología , Triglicéridos/sangreRESUMEN
BACKGROUND: Garlic (Allium sativum) has been considered to exhibit therapeutic features for many years. The effects of garlic on levels of serum lipids and on atherosclerosis have been investigated extensively. We have previously demonstrated that allicin, an active component of garlic, exerts a beneficial effect on lipid profile in hyperlipidemic rabbits. OBJECTIVE: To investigate the effects of allicin on formation of fatty streaks (atherosclerosis) and lipid profile in mice. METHODS: Allicin was extracted from garlic and kept in a buffer citrate solution at 4 degrees C. Sixty C57BL/6 mice were fed Paigen diet (17% fat, 1.25% cholesterol) for 15 weeks. Thirty randomly selected animals were administered allicin solution (9 mg/kg) and 30 were administered placebo. Blood lipid profile was evaluated five times during the study. At the end of the 15-week period, the animals were killed and the aortic sinus was evaluated for formation of fatty streaks (atherosclerosis). RESULTS: We observed no statistically significant differences between blood lipid profiles of groups. Microscopic evaluation of aortic sinus formation of fatty streaks (atherosclerosis), however, showed that values for mice in the allicin-treated group were significantly lower: areas of formation of fatty streaks (atherosclerosis) were 13,440 +/- 3310 and 23,410 +/- 3723 micron 2, respectively, for allicin-treated and control mice (means +/- SEM; P = 0.023). CONCLUSIONS: These results indicate that allicin reduces formation of fatty streaks (atherosclerosis) in hyperlipidemic mice. These changes do not seem to occur through an alteration in blood lipid profile.
Asunto(s)
Arteriosclerosis/patología , Arteriosclerosis/prevención & control , Colesterol en la Dieta/efectos adversos , Hipolipemiantes/farmacología , Ácidos Sulfínicos/farmacología , Animales , Arteriosclerosis/sangre , HDL-Colesterol/sangre , Disulfuros , Femenino , Ratones , Ratones Endogámicos C57BLRESUMEN
Although lipid oxidation plays a major role in atherogenesis, the role of antioxidants in the prevention and treatment of the process is not clear. Apolipoprotein (apo) E-deficient mice develop spontaneous atherosclerotic lesions in major arteries. The presence of oxidized lipoprotein epitopes in the lesion suggests that oxidation reactions are involved in atherogenesis in this mouse model, but the inhibitory effect of antioxidants on atherogenesis in the model is controversial. To test the effect of dietary antioxidants on atherogenesis, male apoE-deficient mice (n=15) were fed a standard chow diet supplemented with 0.05% alpha-tocopherol and 0.05% all-trans beta-carotene. A control group (n=15) received no antioxidant supplement. At the end of the trial, mice consuming vitamins had 5x more plasma vitamin E but undetectable beta-carotene levels. However, liver levels of the beta-carotene metabolite, retinyl palmitate, were higher in antioxidant-treated mice compared with control mice. The antioxidants had no effect on lipoprotein or on plasma anti-oxidatively modified low density lipoproteins (anti-oxLDL) antibody levels. The vitamins had a small but insignificant effect on lipoprotein resistance to ex vivo oxidation, determined by a longer lag period of conjugated diene formation. Atherosclerosis, determined by the lesion size at the aortic sinus, was insignificantly suppressed in antioxidant-treated mice (mean area+/-SE, 20 000+/-7129 versus 13 281+/-5861 micrometer(2); P=0.40). The aortic atherosclerotic lesion area was similar in both experimental groups (2.55+/-0.65% and 2.08+/-0.5% of total aortic area in the control and antioxidant group, respectively; P=0.58). The results of the current study suggest that moderate levels of synthetic antioxidant vitamins have no effect on atherogenesis in apoE-deficient mice.
Asunto(s)
Antioxidantes/farmacología , Apolipoproteínas E/deficiencia , Arteriosclerosis/prevención & control , Vitamina E/administración & dosificación , beta Caroteno/administración & dosificación , Animales , Antioxidantes/análisis , Peso Corporal , Colesterol/sangre , Lipoproteínas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Oxidación-ReducciónRESUMEN
Magnesium, an important cofactor of more than 300 enzymes, has previously been found to modulate blood lipid levels, atherogenesis and atherosclerosis in rabbits, when added to their diet. The aim of this study was to examine whether magnesium fortification of drinking water, without a change in diet content, can affect atherogenesis. The study included six groups of LDL-receptor-deficient mice. The mice received either distilled water or water containing 50 g of magnesium sulfate per liter. In the first (12 weeks) and second (6 weeks) stages of the experiment, the mice received low- and high-cholesterol diets, respectively. At the end of each stage, blood was drawn for the determination of plasma magnesium, calcium and lipid levels. In addition, the extent of atherosclerosis was determined at the aortic sinus. In both males and females, magnesium fortification was associated with higher levels of plasma magnesium (50 and 37% increase, respectively), without any differences in plasma calcium content. The extent of atherosclerosis at the aortic sinus in the male mice that received high levels of magnesium was a third of that of the male mice that received distilled water. However, these differences were not found in the female groups. Surprisingly, the female mice that received water fortified with magnesium had higher levels of cholesterol after stage 2, whereas no differences regarding plasma lipid levels were found among the male mice. These results confirm that magnesium fortification of drinking water is capable of inhibiting atherogenesis in male LDL-receptor-deficient mice. The mechanisms of action are yet to be discovered, and are probably not related to diminished lipid excretion, but possibly to the prevention of calcium influx into vascular smooth muscle cells, elevated antioxidative capacity, or other yet undetermined mechanisms.
Asunto(s)
Arteriosclerosis/prevención & control , Sulfato de Magnesio/administración & dosificación , Receptores de LDL/deficiencia , Animales , Aorta Torácica/patología , Arteriosclerosis/sangre , Arteriosclerosis/patología , Calcio/sangre , Modelos Animales de Enfermedad , Ingestión de Líquidos , Lípidos/sangre , Magnesio/sangre , Masculino , Ratones , Receptores de LDL/sangreRESUMEN
BACKGROUND: Immunization with beta2-glycoprotein I (beta2GPI), the probable target of autoimmune anticardiolipin antibodies, results in experimental antiphospholipid syndrome in different mouse strains. The present study was undertaken to evaluate the effect of beta2GPI immunization on the progression of atherosclerosis. METHODS AND RESULTS: In the first experiment, 3 groups of LDL receptor-deficient (LDL-RD) mice (n=15 per group) were immunized with either beta2GPI or ovalbumin or were not immunized and were fed a chow diet for 12 weeks. In a second experiment, 3 groups of LDL-RD mice (n=10 per group) were immunized similarly and fed an atherogenic diet for 6 weeks. All beta2GPI-immunized mice developed high titers of anti-beta2GPI antibodies as well as a specific lymph node proliferation to beta2GPI. The average cholesterol levels did not differ between the mice fed similar diets, regardless of the immunization protocol. Atherosclerosis was enhanced in the beta2GPI-immunized mice (mean aortic lesion, 26 000+/-5700 microm2) in comparison with their ovalbumin-immunized (mean, 3000+/-1099 microm2; P<0.01) and nonimmunized (mean, 2250+/-700 microm2; P<0.01) littermates. The average lesion size in the beta2GPI-immunized mice fed an atherogenic diet (mean, 98 000+/-8305 microm2) was larger than the ovalbumin-immunized mice (mean, 81 250+/-12 933 microm2; P=NS) or the nonimmunized controls (mean, 75 625+/-7281 microm2; P=NS). The atherosclerotic plaques in the beta2GPI-immunized mice appeared to be more mature, and denser infiltration of CD4 lymphocytes was present in the subendothelium of the aortic sinuses from this group of mice. CONCLUSIONS: The results of the present study provide the first direct evidence for the proatherogenic effect of ss2GPI immunization and establish a new model for immune-mediated atherosclerosis.
Asunto(s)
Arteriosclerosis/inmunología , Proteínas Bacterianas , Glicoproteínas/inmunología , Receptores de LDL/genética , Animales , Anticuerpos/farmacología , Especificidad de Anticuerpos , Complejo Antígeno-Anticuerpo/sangre , Estenosis de la Válvula Aórtica/inmunología , Apolipoproteínas/inmunología , Peso Corporal , Linfocitos T CD4-Positivos/inmunología , Chaperonina 60 , Chaperoninas/inmunología , LDL-Colesterol/sangre , LDL-Colesterol/inmunología , Dieta , Femenino , Inmunización , Inmunohistoquímica , Ganglios Linfáticos/citología , Ganglios Linfáticos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Ovalbúmina/inmunología , beta 2 Glicoproteína IRESUMEN
The role of the immune system in modulating atherosclerosis has recently been the subject of intensive research. Several previous authors have put forward a paradigm of the autoimmune process occurring in the vicinity of the plaque. Two recent studies have shown that immunization of rabbits with homologous modified low-density lipoprotein (LDL) led to suppression of atherosclerosis. In the current study we evaluated the effects of homologous malondialdehyde (MDA)-LDL immunizations on atherogenesis in apo-E-deficient mice. Two groups of female chow-diet-fed, apo-E-deficient mice (n = 10) were either immunized with homologous MDA-LDL or with phosphate buffer saline (PBS) at 2-week intervals. The mice were sacrificed 12 weeks following the primary immunization. The MDA-LDL-immunized mice were shown to develop high titers of anti-MDA-LDL antibodies. Atherosclerosis, determined by the lesion size at the aortic sinus, was significantly suppressed in the MDA-LDL-immunized mice as compared with their littermates immunized with PBS (mean area +/- S.D.; 74000 +/- 17300 microm2 versus 158000 +/- 12800 microm2; P < 0.01). No differences were found between the groups with respect to the cellular composition of the atherosclerotic plaques. The results of this study show that immunization with MDA-LDL has a protective effect in apo-E-deficient mice, and further suggests that this mouse model is suitable for studies of immunomodulation.
Asunto(s)
Apolipoproteínas E/deficiencia , Arteriosclerosis/prevención & control , Inmunización , Lipoproteínas LDL/inmunología , Malondialdehído/inmunología , Animales , Anticuerpos/análisis , Arteriosclerosis/patología , Femenino , Inmunohistoquímica , Lípidos/sangre , Ratones , Seno Aórtico/patologíaRESUMEN
Heparan sulfate and low density lipoprotein receptor related protein (LRP) have been shown to participate in the uptake and degradation of the enzyme lipoprotein lipase (LPL). Yet, the contribution of each of these pathways to LPL metabolism and their possible dependence is unknown. In the present study we examined the metabolism of 125I-labeled LPL in untreated and heparinase-treated primary wild-type mouse embryonic fibroblasts (MEF) and in mouse fibroblasts that express single LRP allele (PEA-10) or are lacking the LRP (PEA-13). The degradation of LPL in PEA-13 cells was 30% lower than in MEF and PEA-10 cells. Heparinase treatment decreased the LPL degradation by 58%, 79% and 92%, whereas heparin reduced such degradation by 87%, 90% and 94% in MEF, PEA-10 and PEA-13 cultures, respectively. Assuming that a) heparinase treatment abolished the heparan-sulfate pathway, and that b) the degradation remaining in heparin-treated cultures represents nonspecific values, it appears that heparan sulfate contributes about 61%, 83% and 95% of total LPL degradation, whereas the LRP pathway contributes 39%, 17% and less than 5% of LPL degradation in MEF, PEA-10 and PEA-13 cells, respectively. In addition, the data indicate that LPL interaction with heparan sulfate and the LRP pathways is independent of each other. The study shows that these cells possess both a heparan sulfate-dependent pathway and an LRP-dependent pathway for LPL metabolism and that the two pathways are independent of each other.
Asunto(s)
Heparitina Sulfato/metabolismo , Lipoproteína Lipasa/metabolismo , Receptores Inmunológicos/metabolismo , Receptores de LDL/metabolismo , Alelos , Animales , Células Cultivadas , Fibroblastos/metabolismo , Heparina/farmacología , Liasa de Heparina , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad , Ratones , Polisacárido Liasas/farmacología , Receptores de LDL/genéticaRESUMEN
Lipoprotein lipase (LPL) is rapidly and efficiently cleared from the circulation by the liver through an as yet unclear mechanism. In the present study, we determined the nature of LPL interactions with the liver parenchimal cell line HepG2 as compared to other cells in culture. Binding, cell association and degradation of 125I-labelled bovine milk LPL by HepG2 cells, normal and low density lipoprotein (LDL) receptor-negative human fibroblasts and Chinese hamster ovary (CHO) cells show similar values irrespective of source and origin. LPL metabolism in HepG2 cells was characterized by a high capacity to degrade the enzyme, an extremely high sensitivity to heparin and was inhibited by 60%-70% after treatment of the cells with sodium chlorate and heparinase (but not chondroitinase). These findings suggested an important role for heparan sulfate in the process of cell interaction and metabolism of LPL. To further clarify the role of heparan sulfate in determining the LPL-cell interactions, we compared the metabolism of LPL in wild type and mutant heparan sulfate-deficient CHO cells. Heparan sulfate-deficient CHO cells show a low capacity to bind and degrade LPL, about 10%-20% that of the wild type cells. In another set of experiments, we sought to determine whether LPL interactions with HepG2 cells are affected by triglyceride-rich lipoproteins. The results clearly show that whereas unlabeled LPL dramatically enhanced the metabolism of radioiodinated very low density lipoprotein (VLDL), unlabeled VLDL had no effect on radioiodinated LPL metabolism in these cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Heparina/farmacología , Heparitina Sulfato/metabolismo , Lipoproteína Lipasa/metabolismo , Animales , Células CHO , Bovinos , Línea Celular , Cricetinae , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Lipoproteína Lipasa/efectos de los fármacos , Lipoproteínas VLDL/efectos de los fármacos , Lipoproteínas VLDL/metabolismo , Hígado/citología , Hígado/metabolismo , Unión ProteicaRESUMEN
To test the possibility that HMG-CoA reductase inhibitors reduce LDL mass by an increased VLDL catabolism, we determined the effect of simvastatin therapy on cellular metabolism of VLDL in 18 patients with primary hypercholesterolaemia. Six months of simvastatin therapy was followed by 26%, 31% and 21% reduction of plasma total cholesterol, LDL-cholesterol and plasma triglyceride levels, respectively. Before therapy, patients' VLDL metabolism in cultured human normal skin fibroblasts was similar to control VLDL. Six months after therapy was initiated, a remarkable 2-5-fold increase in VLDL cell metabolism was found. These effects were even more marked when the VLDL was enriched with exogenous recombinant apo E-3. A comparison of the metabolism of the patients' VLDL to control VLDL and LDL, revealed that simvastatin increased metabolic ratios of 60-70% and 45-95%, respectively. Simvastatin therapy was associated with a decrease of VLDL cholesteryl ester content of 19% and increase of the phospholipid content of 13%. The data strongly indicate that simvastatin therapy stimulates VLDL: cell interactions and catabolism, possibly reflecting alterations of the physico-chemical properties of the particle. It is proposed that in addition to other previously described pathways, HMG-CoA reductase inhibitors decrease LDL mass through a novel mechanism of enhanced VLDL catabolism prior to the conversion to LDL.
Asunto(s)
Anticolesterolemiantes/farmacología , Hidroximetilglutaril-CoA Reductasas/análisis , Lipoproteínas VLDL/metabolismo , Lovastatina/análogos & derivados , Adulto , Anciano , Femenino , Humanos , Lipoproteínas LDL/metabolismo , Lovastatina/farmacología , Masculino , Persona de Mediana Edad , SimvastatinaAsunto(s)
Adenilil Ciclasas/metabolismo , Nucleótidos de Guanina/análogos & derivados , Nucleótidos de Guanina/farmacología , Guanosina Difosfato/análogos & derivados , Guanosina Difosfato/farmacología , Guanosina Trifosfato/farmacología , Tionucleótidos/farmacología , Animales , Membrana Celular/enzimología , Activación Enzimática/efectos de los fármacos , Membrana Eritrocítica/enzimología , Fluoruros/farmacología , Guanilil Imidodifosfato/farmacología , Isoproterenol/farmacología , Cinética , Hígado/enzimología , Glándula Parótida/enzimología , Propranolol/farmacología , Ratas , PavosRESUMEN
It has recently been suggested that adenylate cyclase activity is controlled by a regulatory cycle consisting of two reactions: a hormone induced formation of the active adenylate cyclase-GTP complex, and a subsequent turn-off reaction in which hydrolysis of the bound nucleotide reverts the system to the inactive state. To test this model each of the two reactions was measured separately and their rate constants were used to estimate the steady state adenylate cyclase and GTPase activities. The first order rate constants were kon = 3 min-1 for the activation reaction and koff = 15 min-1 for the turn-off reaction. Substitution of these rate constants in the steady state equation of the regulatory cycle gave values of hormone stimulated adenylate cyclase and GTPase activities similar to those determined by direct measurements. Treatment of the adenylate cyclase with cholera toxin caused a decrease of 96% in the rate constant of the turn-off reaction. In this case too the activities calculated from the steady state equation were in good agreement with those determined directly.